Project description:Raw data files (RAW and mzML) and Scaffold search engine files corresponding to the whole proteome analyses of the syphilis spirochete, Treponema pallidum ssp. pallidum, strain SS14.
Project description:Proteome-wide analysis of the syphilis spirochete, Treponema pallidum ssp. pallidum (Nichols strain). Treponemes were in vitro-cultured.
Project description:Proteome-wide analysis of the syphilis spirochete, Treponema pallidum ssp. pallidum (Nichols strain). Treponemes were cultured in, and isolated from, New Zealand white rabbits.
Project description:Two new tprD alleles have been identified in Treponema pallidum: tprD2 is found in 7 of 12 T. pallidum subsp. pallidum isolates and 7 of 8 non-pallidum isolates, and tprD3 is found in one T. pallidum subsp. pertenue isolate. Antibodies against TprD2 are found in persons with syphilis, demonstrating that tprD2 is expressed during infection.
Project description:Pathogens adapt and evolve in response to pressures exerted by host environments, leading to generation of genetically diverse variants. Treponema pallidum subspecies pallidum displays a substantial amount of interstrain diversity. These variants have been identified in various parts of the world, indicating transmission linkage between geographical regions. Genotyping is based on molecular characterisation of various loci in the syphilis treponeme genome, but still require further development and continued research, as new bacterial types are continually being detected. The goal for studying the molecular epidemiology of Treponema pallidum variants is the global monitoring of the transmission of genetically distinct organisms with different drug sensitivities and, potentially, different virulence proprieties.
Project description:We performed a comprehensive miRNA profiling analysis of exosomes by Treponema pallidum-stimulated microarrays. A total of 2×106 macrophages were obtained by THP-1 differentiation and grown in RPMI-1640 containing 10% exosome-free FBS. Exosomes were acquired from macrophage culture supernatants with (n = 7) or without (n = 3) T. pallidum. Briefly, macrophages were washed in PBS twice and further grown in fresh medium for 12 h (n = 2), 24 h (n = 2) and 48 h (n = 3) to collect exosomes. Exosomal miRNA microarray assays were carried out with Agilent Human miRNA (8*60K) array.
Project description:The outer membrane (OM) of Treponema pallidum, the uncultivatable agent of venereal syphilis, has long been the subject of misconceptions and controversy. Decades ago, researchers postulated that T. pallidum's poor surface antigenicity is the basis for its ability to cause persistent infection, but they mistakenly attributed this enigmatic property to the presence of a protective outer coat of serum proteins and mucopolysaccharides. Subsequent studies revealed that the OM is the barrier to antibody binding, that it contains a paucity of integral membrane proteins, and that the preponderance of the spirochete's immunogenic lipoproteins is periplasmic. Since the advent of recombinant DNA technology, the fragility of the OM, its low protein content, and the lack of sequence relatedness between T. pallidum and Gram-negative outer membrane proteins (OMPs) have complicated efforts to characterize molecules residing at the host-pathogen interface. We have overcome these hurdles using the genomic sequence in concert with computational tools to identify proteins predicted to form β-barrels, the hallmark conformation of OMPs in double-membrane organisms and evolutionarily related eukaryotic organelles. We also have employed diverse methodologies to confirm that some candidate OMPs do, in fact, form amphiphilic β-barrels and are surface-exposed in T. pallidum. These studies have led to a structural homology model for BamA and established the bipartite topology of the T. pallidum repeat (Tpr) family of proteins. Recent bioinformatics has identified several structural orthologs for well-characterized Gram-negative OMPs, suggesting that the T. pallidum OMP repertoire is more Gram-negative-like than previously supposed. Lipoprotein adhesins and proteases on the spirochete surface also may contribute to disease pathogenesis and protective immunity.
Project description:Despite more than a century of research, genetic manipulation of Treponema pallidum subsp. pallidum (T. pallidum), the causative agent of syphilis, has not been successful. The lack of genetic engineering tools has severely limited understanding of the mechanisms behind T. pallidum success as a pathogen. A recently described method for in vitro cultivation of T. pallidum, however, has made it possible to experiment with transformation and selection protocols in this pathogen. Here, we describe an approach that successfully replaced the tprA (tp0009) pseudogene in the SS14 T. pallidum strain with a kanamycin resistance (kanR) cassette. A suicide vector was constructed using the pUC57 plasmid backbone. In the vector, the kanR gene was cloned downstream of the tp0574 gene promoter. The tp0574prom-kanR cassette was then placed between two 1-kbp homology arms identical to the sequences upstream and downstream of the tprA pseudogene. To induce homologous recombination and integration of the kanR cassette into the T. pallidum chromosome, in vitro-cultured SS14 strain spirochetes were exposed to the engineered vector in a CaCl2-based transformation buffer and let recover for 24 hours before adding kanamycin-containing selective media. Integration of the kanR cassette was demonstrated by qualitative PCR, droplet digital PCR (ddPCR), and whole-genome sequencing (WGS) of transformed treponemes propagated in vitro and/or in vivo. ddPCR analysis of RNA and mass spectrometry confirmed expression of the kanR message and protein in treponemes propagated in vitro. Moreover, tprA knockout (tprAko-SS14) treponemes grew in kanamycin concentrations that were 64 times higher than the MIC for the wild-type SS14 (wt-SS14) strain and in infected rabbits treated with kanamycin. We demonstrated that genetic manipulation of T. pallidum is attainable. This discovery will allow the application of functional genetics techniques to study syphilis pathogenesis and improve syphilis vaccine development.
Project description:BackgroundCurrent syphilis tests cannot distinguish between active and past syphilis among patients with serofast rapid plasma reagin (RPR) titers. We investigated whether cytokine profiles might provide insight in the differentiation of active and treated syphilis.MethodsWe collected quarterly serum samples from participants at risk for incident syphilis in a prospective cohort study of men and male-to-female transgender women. We defined incident syphilis as a new RPR titer ≥ 1:8 or a fourfold increase from a prior RPR titer and a positive Treponema pallidum particle agglutination assay. We measured cytokine expression using a 63-multiplex bead-based Luminex assay (eBiosciences/Affymetrix, San Diego, California, USA). We used tertile bins and Chi square tests to identify differences in proportions of cytokines between samples from patients with active and treated syphilis. We constructed a network of cytokine profiles from those findings. We used R software (R version 3.4.1, R, Vienna, Austria) to fit models.ResultsWe identified 20 pairs of cytokines (out of 1953 possible pairs) that differed between active and treated syphilis. From those, we identified three cytokine networks of interest: an Eotaxin-Rantes-Leptin network, a Mig-IL1ra-Trail-CD40L network, and an IL12p40-IL12p70 network.ConclusionsDifferences in cytokine profiles are present among men and male-to-female transgender women with active and treated syphilis. Cytokine assays may be a potentially useful tool for identifying active syphilis among patients with serologic syphilis reactivity.